Discovery Of Small Molecule Inhibitors Research Articles

Molecular Modeling Insights into Upadacitinib Selectivity upon Binding to JAK Protein Family.

Rheumatoid arthritis (RA) is a chronic disease characterized by bone joint damage and incapacitation. The mechanism underlying RA pathogenesis is autoimmunity in the connective tissue. Cytokines play an important role in the human immune system for signal transduction and in the development of inflammatory responses. Janus kinases (JAK) participate in the JAK/STAT pathway, which mediates cytokine effects, in particular interleukin 6 and IFNγ. The discovery of small molecule inhibitors of the JAK protein family has led to a revolution in RA therapy. The novel JAK inhibitor upadacitinib (RinvoqTM) has a higher selectivity for JAK1 compared to JAK2 and JAK3 in vivo. Currently, details on the molecular recognition of JAK1 by upadacitinib are not available. We found that characteristics of hydrogen bond formation with the glycine loop and hinge in JAKs define the selectivity. Our molecular modeling study could provide insight into the drug action mechanism and pharmacophore model differences in JAK isoforms.

Structural effects of morpholine replacement in ZSTK474 on Class I PI3K isoform inhibition: Development of novel MEK/PI3K bifunctional inhibitors

Established roles for PI3K and MAPK signaling pathways in tumorigenesis has prompted extensive research towards the discovery of small-molecule inhibitors as cancer therapeutics. However, significant compensatory regulation exists between these two signaling cascades, leading to redundancy among survival pathways. Consequently, initial clinical trials aimed at either PI3K or MEK inhibition alone have proven ineffective and highlight the need for development of targeted and innovative therapeutic combination strategies. We designed a series of PI3K inhibitor derivatives wherein a single morpholine group of the PI3K inhibitor ZSTK474 was substituted with a variety of 2-aminoethyl functional groups. Analogs with pendant hydroxyl or methoxy groups maintained low nanomolar inhibition towards PI3Kα, PI3Kγ, and PI3Kδ isoforms in contrast to those with pendant amino groups which were significantly less inhibitory. Synthesis of prototype PI3K/MEK bifunctional inhibitors (6r, 6s) was guided by the structure-activity data, where a MEK-targeting inhibitor was tethered directly via a short PEG linker to the triazine core of the PI3K inhibitor analogs. These compounds (6r, 6s) displayed nanomolar inhibition towards PI3Kα, δ, and MEK (IC50 ∼105–350 nM), and low micromolar inhibition for PI3Kβ and PI3Kγ (IC50 ∼1.5–3.9 μM) in enzymatic inhibition assays. Cell viability assays demonstrated superior anti-proliferative activity for 6s over 6r in three tumor-derived cell lines (A375, D54, SET-2), which correlated with inhibition of downstream AKT and ERK1/2 phosphorylation. Compounds 6r and 6s also demonstrated in vivo tolerability with therapeutic efficacy through reduction of kinase activation and amelioration of disease phenotypes in the JAK2V617F mutant myelofibrosis mouse cancer model. Taken together, these results support further structure optimization of 6r and 6s as promising leads for combination therapy in human cancer as a new class of PI3K/MEK bifunctional inhibitors.

Fragment-Based Discovery of Small Molecules Bound to T-Cell Immunoglobulin and Mucin Domain-Containing Molecule 3 (TIM-3).

T-cell immunoglobulin and mucin domain-containing molecule 3 (TIM-3; HAVCR2) has emerged as an attractive immune checkpoint target for cancer immunotherapy. TIM-3 is a negative regulator of the systemic immune response to cancer and is expressed on several dysfunctional, or exhausted, immune cell subsets. Upregulation of TIM-3 is associated with tumor progression, poor survival rates, and acquired resistance to antibody-based immunotherapies in the clinic. Despite the potential advantages of small-molecule inhibitors over antibodies, the discovery of small-molecule inhibitors has lagged behind that of antibody therapeutics. Here, we describe the discovery of high-affinity small-molecule ligands for TIM-3 through an NMR-based fragment screen and structure-based lead optimization. These compounds represent useful tools to further study the biology of TIM-3 immune modulation in cancer and serve as a potentially useful starting point toward the discovery of TIM-3-targeted therapeutics.

Glioma-Targeted Therapeutics: Computer-Aided Drug Design Prospective.

Amongst the several types of brain cancers known to humankind, glioma is one of the most severe and life-threatening types of cancer, comprising 40% of all primary brain tumors. Recent reports have shown the incident rate of gliomas to be 6 per 100,000 individuals per year globally. Despite the various therapeutics used in the treatment of glioma, patient survival rate remains at a median of 15months after undergoing first-line treatment including surgery, radiation, and chemotherapy with Temozolomide. As such, the discovery of newer and more effective therapeutic agents is imperative for patient survival rate. The advent of computer-aided drug design in the development of drug discovery has emerged as a powerful means to ascertain potential hit compounds with distinctively high therapeutic effectiveness against glioma. This review encompasses the recent advances of bio-computational in-silico modeling that have elicited the discovery of small molecule inhibitors and/or drugs against various therapeutic targets in glioma. The relevant information provided in this report will assist researchers, especially in the drug design domains, to develop more effective therapeutics against this global disease.

Valosin-Containing Protein (VCP)/p97: A Prognostic Biomarker and Therapeutic Target in Cancer.

Valosin-containing protein (VCP)/p97, a member of the AAA+ ATPase family, is a molecular chaperone recruited to the endoplasmic reticulum (ER) membrane by binding to membrane adapters (nuclear protein localization protein 4 (NPL4), p47 and ubiquitin regulatory X (UBX) domain-containing protein 1 (UBXD1)), where it is involved in ER-associated protein degradation (ERAD). However, VCP/p97 interacts with many cofactors to participate in different cellular processes that are critical for cancer cell survival and aggressiveness. Indeed, VCP/p97 is reported to be overexpressed in many cancer types and is considered a potential cancer biomarker and therapeutic target. This review summarizes the role of VCP/p97 in different cancers and the advances in the discovery of small-molecule inhibitors with therapeutic potential, focusing on the challenges associated with cancer-related VCP mutations in the mechanisms of resistance to inhibitors.

Collagen I Induces Preeclampsia-Like Symptoms by Suppressing Proliferation and Invasion of Trophoblasts.

Preeclampsia is a common obstetric disorder affecting 2-8% of pregnancy worldwide. Fibrosis is an important histological change occurring in preeclamptic placenta, and might depend on the excess deposition of collagen I. However, the role of fibrotic placenta and collagen I in the pathogenesis of preeclampsia remains unclear. Therefore, we analyzed the collagen deposition and the expression of Collagen I in human placenta by Masson staining, Sirius red staining and western blotting. Further, the role of collagen I in preeclampsia pathogenesis was studied in C57BL/6 mice. HTR-8/SVneo cells were used to investigate the mechanisms underlying the effects of collagen I in trophoblasts by transcriptome sequencing and pharmacological agonists. Human preeclamptic placenta exhibited a significantly higher degree of fibrosis in stem villi and terminal villi than normal placenta, and was characterized by collagen I deposition. In vivo, a single injection of collagen I on gestational day 0.5 led to an increase in systolic pressure of pregnant mice from gestational days 4.5–17.5, to a decrease in weight and number of embryos, and to enhanced placental collagen I expression and degree of fibrosis compared with control mice. In vitro, collagen I attenuated the proliferation and invasion of HTR-8SV/neo cells. This effect could be reversed by treatment with agonists of ERK and β-catenin. Moreover, transcriptome sequencing demonstrated that signaling pathways related to cell proliferation and invasion were significantly downregulated in HTR-8SV/neo cells. Thus, we propose that collagen I induced preeclampsia-like symptoms by suppressing the proliferation and invasion of trophoblasts through inhibition of the ERK phosphorylation and WNT/β-catenin signaling pathways. Our findings could pave the way to the discovery of small-molecule inhibitors for preeclampsia treatment and future studies with larger sample size are required.

Proteases and Their Modulators in Cancer Therapy: Challenges and Opportunities.

Proteostasis is the process of regulating intracellular proteins to maintain the balance of the cell proteome, which is crucial for cancer cell survival. Several proteases located in the cytoplasm, mitochondria, lysosome, and extracellular environment have been identified as potential antitumor targets because of their involvement in proteostasis. Although the discovery of small-molecule inhibitors targeting proteases faces particular challenges, rapid advances in chemical biology and structural biology, and the new technology of drug discovery have facilitated the development of promising protease modulators. In this review, the protein structure and function of important tumor-related proteases and their inhibitors are presented. We also provide a prospective on advances and the outlook of new drug strategies that target these proteases.

Identification and characterization of second-generation EZH2 inhibitors with extended residence times and improved biological activity

The histone methyltransferase EZH2 has been the target of numerous small-molecule inhibitor discovery efforts over the last 10+ years. Emerging clinical data have provided early evidence for single agent activity with acceptable safety profiles for first-generation inhibitors. We have developed kinetic methodologies for studying EZH2-inhibitor-binding kinetics that have allowed us to identify a unique structural modification that results in significant increases in the drug-target residence times of all EZH2 inhibitor scaffolds we have studied. The unexpected residence time enhancement bestowed by this modification has enabled us to create a series of second-generation EZH2 inhibitors with sub-pM binding affinities. We provide both biophysical evidence validating this sub-pM potency and biological evidence demonstrating the utility and relevance of such high-affinity interactions with EZH2.

Discovery of small-molecule inhibitors of multidrug-resistance plasmid maintenance using a high-throughput screening approach

Carbapenem-resistant Enterobacteriaceae (CRE) are multidrug-resistant pathogens for which new treatments are desperately needed. Carbapenemases and other types of antibiotic resistance genes are carried almost exclusively on large, low-copy-number plasmids (pCRE). Accordingly, small molecules that efficiently evict pCRE plasmids should restore much-needed treatment options. We therefore designed a high-throughput screen to identify such compounds. A synthetic plasmid was constructed containing the plasmid replication machinery from a representative Escherichia coli CRE isolate as well as a fluorescent reporter gene to easily monitor plasmid maintenance. The synthetic plasmid was then introduced into an E. coli K12 tolC host. We used this screening strain to test a library of over 12,000 known bioactive agents for molecules that selectively reduce plasmid levels relative to effects on bacterial growth. From 366 screen hits we further validated the antiplasmid activity of kasugamycin, an aminoglycoside; CGS 15943, a nucleoside analog; and Ro 90-7501, a bibenzimidazole. All three compounds exhibited significant antiplasmid activity including up to complete suppression of plasmid replication and/or plasmid eviction in multiple orthogonal readouts and potentiated activity of the carbapenem, meropenem, against a strain carrying the large, pCRE plasmid from which we constructed the synthetic screening plasmid. Additionally, we found kasugamycin and CGS 15943 blocked plasmid replication, respectively, by inhibiting expression or function of the plasmid replication initiation protein, RepE. In summary, we validated our approach to identify compounds that alter plasmid maintenance, confer resensitization to antimicrobials, and have specific mechanisms of action.

The Discovery of Insulin-Regulated Aminopeptidase (IRAP) Inhibitors: A Literature Review.

Insulin-Regulated Aminopeptidase (IRAP, EC 3.4.11.3) is a multi-tasking member of the M1 family of zinc aminopeptidases. Among its diverse biological functions, IRAP is a regulator of oxytocin levels during late stages of pregnancy, it affects cellular glucose uptake by trafficking of the glucose transporter type 4 and it mediates antigen cross-presentation by dendritic cells. Accumulating evidence show that pharmacological inhibition of IRAP may hold promise as a valid approach for the treatment of several pathological states such as memory disorders, neurodegenerative diseases, etc. Aiming to the investigation of physiological roles of IRAP and therapeutic potential of its regulation, intense research efforts have been dedicated to the discovery of small-molecule inhibitors. Moreover, reliable structure-activity relationships have been largely facilitated by recent crystal structures of IRAP and detailed computational studies. This review aims to summarize efforts of medicinal chemists toward the design and development of IRAP inhibitors, with special emphasis to factors affecting inhibitor selectivity.

Conformational Selection in Ligand Recognition by the First Tudor Domain of PHF20L1.

The first Tudor domain (Tudor1) of PHF20L1 recognizes (non)histone methylation to play versatile roles. However, the underlying ligand-recognition mechanism remains unknown as a closed state revealed in the free-form structure. NMR relaxation dispersion and molecular dynamics simulations suggest a pre-existing low-population conformation with a remarkable rearrangement of aromatic cage residues of PHF20L1 Tudor1. Such an open-form conformation is utilized to recognize lysine 142 methylated DNMT1, a cosolvent, and an NMR fragment screening hit, as revealed by the complex crystal structures. Intriguingly, the ligand binding capacity was enhanced by mutation that tunes up the open-state population only. The recognition of DNMT1 by PHF20L1 was further validated in cancer cells. This conformational selection mechanism will enable the discovery of small molecule inhibitors against the seemingly "undruggable" PHF20L1 Tudor1.

Discovery of small molecule inhibitors against the NS3/4A serine protease of Hepatitis C virus genotype 3 via highthroughput virtual screening and in vitro evaluations

The hepatitis C virus (HCV) consists of eight genotypes and 90 subtypes, with genotype (GT) 3 being the second most common globally and is linked to higher incidences of steatosis and rapid development of fibrosis and cirrhosis. The NS3/4A serine protease, a heterodimer complex of two HCV non-structural proteins, is an effective target for pharmaceutical intervention due to its essential roles in processing HCV polyproteins and inhibiting innate immunity. This study combines structure-based virtual screening (SBVS) of predefined compound libraries, pharmacokinetic prediction (ADME/T) and in vitro evaluation to identify potential low molecular weight (<500 Dalton) inhibitors of the NS3/4A serine protease (GT3). In silico screening of ZINC and PubChem libraries yielded five selected compounds as potential candidates. Dose-dependent inhibition of the NS3/4A serine protease and HCV replication in HuH-7.5 cells revealed that compound A (PubChem ID No. 16672637) exhibited inhibition towards HCV GT3 with an IC50 of 106.7µM and EC50 of 25.8µM, respectively. Thus, compound A may be developed as a potent, low molecular weight drug against the HCV NS3/4A serine protease of GT3.

Discovery of Small Molecule Inhibitors of Huntingtin Exon 1 Aggregation by FRET-Based High-Throughput Screening in Living Cells.

Huntington's disease (HD) is the most common inherited neurodegenerative disorder and one of the nine polyglutamine (polyQ) diseases. HD is characterized by the pathological aggregation of the misfolded huntingtin exon 1 protein (Httex1) with abnormally long polyQ expansion due to genetic mutation. While there is currently no effective treatment for HD, inhibition of aggregate formation represents a direct approach in mediating the toxicity associated with Httex1 misfolding. To exploit this therapeutic window, we engineered two fluorescence resonance energy transfer (FRET) based biosensors that monitor the aggregation of Httex1 with different expanded Q-lengths (Q39 and Q72) in living cells. These FRET biosensors, together with a high-precision fluorescence lifetime detection platform, enable high-throughput screening of small molecules that target Httex1 aggregation. We found six small molecules that decreased the FRET of the biosensors and reduced Httex1-Q72-induced neuronal cytotoxicity in N2a cells with nanomolar potency. Using advanced SPR and EPR techniques, we confirmed that the compounds directly bind to Httex1 fibrils and inhibit aggregate formation. This strategy in targeting the Httex1 aggregates can be applicable to other proteins involved in polyQ related diseases.

Abstract 14: Computer-aided discovery of small-molecule inhibitors targeting neural transcription factor BRN2 in neuroendocrine prostate tumors

Abstract Therapies targeting tumor-fueling androgens have been mainstay treatments of advanced prostate cancer for almost 5 decades. While the androgen receptor (AR) inhibitor enzalutamide (ENZ) prolongs survival of castration-resistant prostate cancer patients (CRPC), ENZ-resistant (ENZR) tumors rapidly recur. Our laboratory has identified that ENZ resistance can occur both in the presence and absence of continued classical AR activity. Lack of AR activity is clinically relevant, as it has been estimated that up to 25% of men who die from advanced CRPC have non-AR driven disease and lethal AR inactive neuroendocrine and anaplastic phenotypes of CRPC emerge after hormone therapy. Recently our group identified the neural transcription factor BRN2 as a major clinically relevant driver of neuroendocrine prostate cancer (NEPC). Hence, using the integrated power of computational drug discovery platform and biologic testing, we identified first-in-field inhibitors for BRN2. First, a homology model of BRN2 protein was generated and used as an input to perform large-scale virtual screening and virtual hits were tested in a series of experiments. Cpd 18-14, a lead BRN2 inhibitor, exhibited profound inhibition of BRN2 in transcriptional assay and binding to the BRN2 in cellular and in vitro assays. Furthermore, Cpd18-14 reduced the expression of neuroendocrine genes SOX2 and NCAM1. Importantly, Cpd18-14 displayed antiproliferative activity specifically in patient-derived BRN2hi NCI-H660 and in house-developed 42DENZRcells while displaying no effect on BRN2low 16DCRPC and LNCaP cells. Further, the most stable binding orientation of Cpd18-14 was determined using molecular dynamics simulations and was used to carry out structure-based lead optimization. As a result, we developed several synthetic derivatives with significant improvement in antiproliferative activity and metabolic stability. Finally, Cpd18-94 reduced the growth of NCI-H660 and 42DENZR NEPC tumors significantly in xenograft models. We anticipate that the developed drug prototypes will lay a foundation for the development of small-molecule therapies capable of combating highly aggressive and lethal form of neuroendocrine tumors. Citation Format: Ravi Munuganti, Daksh Thaper, Adeleke Aguda, Soojin Kim, Olena Sivak, Amina Zoubeidi. Computer-aided discovery of small-molecule inhibitors targeting neural transcription factor BRN2 in neuroendocrine prostate tumors [abstract]. In: Proceedings of the AACR Special Conference on Advancing Precision Medicine Drug Development: Incorporation of Real-World Data and Other Novel Strategies; Jan 9-12, 2020; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_1):Abstract nr 14.

Optimization of High-Throughput Methyltransferase Assays for the Discovery of Small Molecule Inhibitors.

Methyltransferases (MTases) play diverse roles in cellular processes. Aberrant methylation levels have been implicated in many diseases, indicating the need for the identification and development of small molecule inhibitors for each MTase. Specific inhibitors can serve as probes to investigate the function and validate therapeutic potential for the respective MTase. High-throughput screening (HTS) is a powerful method to identify initial hits for further optimization. Here, we report the development of a fluorescence-based MTase assay and compare this format with the recently developed MTase-Glo luminescence assay for application in HTS. Using protein N-terminal methyltransferase 1 (NTMT1) as a model system, we miniaturized to 1536-well quantitative HTS format. Through a pilot screen of 1428 pharmacologically active compounds and subsequent validation, we discovered that MTase-Glo produced lower false positive rates than the fluorescence-based MTase assay. Nevertheless, both assays displayed robust performance along with low reagent requirements and can potentially be employed as general HTS formats for the discovery of inhibitors for any MTase.

Aspartate/asparagine-\u03b2-hydroxylase: a high-throughput mass spectrometric assay for discovery of small molecule inhibitors

The human 2-oxoglutarate dependent oxygenase aspartate/asparagine-β-hydroxylase (AspH) catalyses the hydroxylation of Asp/Asn-residues in epidermal growth factor-like domains (EGFDs). AspH is upregulated on the surface of malign cancer cells; increased AspH levels correlate with tumour invasiveness. Due to a lack of efficient assays to monitor the activity of isolated AspH, there are few reports of studies aimed at identifying small-molecule AspH inhibitors. Recently, it was reported that AspH substrates have a non-canonical EGFD disulfide pattern. Here we report that a stable synthetic thioether mimic of AspH substrates can be employed in solid phase extraction mass spectrometry based high-throughput AspH inhibition assays which are of excellent robustness, as indicated by high Z’-factors and good signal-to-noise/background ratios. The AspH inhibition assay was applied to screen approximately 1500 bioactive small-molecules, including natural products and active pharmaceutical ingredients of approved human therapeutics. Potent AspH inhibitors were identified from both compound classes. Our AspH inhibition assay should enable the development of potent and selective small-molecule AspH inhibitors and contribute towards the development of safer inhibitors for other 2OG oxygenases, e.g. screens of the hypoxia-inducible factor prolyl-hydroxylase inhibitors revealed that vadadustat inhibits AspH with moderate potency.

Discovery of small molecule inhibitors through pharmacophore modeling, molecular docking, molecular dynamics simulation and experimental validation against myeloid cell leukemia-1 (Mcl-1)

Myeloid cell leukemia-1 (Mcl-1) protein is a family of Bcl-2 (B cell lymphoma 2) rich proteases of the most common increase threshold for genetic aberrations observed in human cancer, including lung, breast, pancreatic, cervical, and ovarian cancers as well as leukemia and lymphoma. Mcl-1 is recognized as an attractive drug target in number of diseases, including cancer. In the present study we surveyed and collected queries compounds from PDB database of Mcl-1 protein and generated pharmacophore-based models adapted to screen the drug-like compounds from FDA approved database. The 206 best lead molecules from pharmacophore-screening were further evaluated by molecular docking, molecular dynamics simulation, MM-GBSA calculation, as well as experimental validation. Two hits, ZINC00601272 and ZINC00002166, showed the best docking scores, which showed a tendency to inhibit cell viability of HL60 and K562 leukemia cells with Mcl-1 expressions. Conclusively, the present study provides structural information of Mcl-1 inhibitors for next generations of cancer therapeutics through computational and experimental validation approach. Communicated by Ramaswamy H. Sarma

Discovery of small molecule inhibitors of chikungunya virus proteins (nsP2 and E1) using in silico approaches

Chikungunya virus (CHIKV) has emerged as a major viral threat, affecting over a million people worldwide per year. It is a vector borne disease transmitted to the human by Ades mosquitoes and primarily affect people by causing viral fever, severe joint pain and other symptoms, like rash, joint swelling, muscle pain and in rare cases can be fatal. CHIKV is a deadly virus, with its mutation rate found to be significantly higher as compared to other viruses. To date, there has been no reported FDA approved drug against this virus. Thus, keeping in mind the urgent need to scrutinize potential therapies against CHIKV, the present study identified twenty plant bioactive compounds that are available at low price and do not have associated adverse effect. For identification of active potentials molecules the pharmacoinformatics-based perspective was applied against CHIKV structural (E1) and non-structural (nsP2) proteins using molecular docking and scoring. The selected compounds were further studied for pharmacokinetics (PK) and pharmacodynamics (PD) associated parameters such as initial absorption, then distribution and later on metabolism excretion and toxicity (ADMET) profiles based on in silico study. The results reveal five potential lead compounds having high binding energy that can help in the development of commercial drugs with favorable ADMET characteristic. Communicated by Ramaswamy H. Sarma

Zoning in on Tankyrases: A Brief Review on the Past, Present and Prospective Studies.

Tankyrases are known for their multifunctionalities within the poly(ADPribose) polymerases family and playing vital roles in various cellular processes which include the regulation of tumour suppressors. Tankyrases, which exist in two isoforms; Tankyrase 1 and 2, are highly homologous and an integral part of the Wnt β -catenin pathway that becomes overly dysregulated when hijacked by pro-carcinogenic machineries. In this review, we cover the distinct roles of the Tankyrase isoforms and their involvement in the disease pathogenesis. Also, we provide updates on experimentally and computationally derived antagonists of Tankyrase whilst highlighting the precedence of integrative computer-aided drug design methods towards the discovery of selective inhibitors. Despite the high prospects embedded in the therapeutic targeting and blockade of Tankyrase isoforms, the inability of small molecule inhibitors to achieve selective targeting has remained a major setback, even until date. This explains numerous incessant drug design efforts geared towards the development of highly selective inhibitors of the respective Tankyrase isoforms since they mediate distinct aberrancies in disease progression. Therefore, considering the setbacks of conventional drug design methods, can computer-aided approaches actually save the day? The implementation of computer-aided drug design techniques in Tankyrase research could help complement experimental methods and facilitate ligand/structure-based design and discovery of small molecule inhibitors with enhanced selectivity.

Computer-Aided Drug Design of β-Secretase, γ-Secretase and Anti-Tau Inhibitors for the Discovery of Novel Alzheimer's Therapeutics.

Aging-associated neurodegenerative diseases, which are characterized by progressive neuronal death and synapses loss in human brain, are rapidly growing affecting millions of people globally. Alzheimer’s is the most common neurodegenerative disease and it can be caused by genetic and environmental risk factors. This review describes the amyloid-β and Tau hypotheses leading to amyloid plaques and neurofibrillary tangles, respectively which are the predominant pathways for the development of anti-Alzheimer’s small molecule inhibitors. The function and structure of the druggable targets of these two pathways including β-secretase, γ-secretase, and Tau are discussed in this review article. Computer-Aided Drug Design including computational structure-based design and ligand-based design have been employed successfully to develop inhibitors for biomolecular targets involved in Alzheimer’s. The application of computational molecular modeling for the discovery of small molecule inhibitors and modulators for β-secretase and γ-secretase is summarized. Examples of computational approaches employed for the development of anti-amyloid aggregation and anti-Tau phosphorylation, proteolysis and aggregation inhibitors are also reported.